title: Ultrarapid generation of femtoliter microfluidic droplets for single-molecule-counting immunoassays.
abstract: We report a microfluidic droplet-based approach enabling the measurement of chemical reactions of individual enzyme molecules and its application to a single-molecule-counting immunoassay. A microfluidic device is used to generate and manipulate <10 fL droplets at rates of up to 1.3 × 10(6) per second, about 2 orders of magnitude faster than has previously been reported. The femtodroplets produced with this device can be used to encapsulate single biomolecular complexes tagged with a reporter enzyme; their small volume enables the fluorescent product of a single enzyme molecule to be detected within 10 min of on-chip incubation. Our prototype system is validated by detection of a biomarker for prostate cancer in buffer, down to a concentration of 46 fM. This work demonstrates a highly flexible and sensitive diagnostic platform that exploits extremely high-speed generation of monodisperse femtoliter droplets for the counting of individual analyte molecules.
authors.affiliation: Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, UK, CB2 1EW. junguk.shim@glasgow.ac.uk
authors.forename: Jung-uk
authors.initials: JU
authors.lastname: Shim
authors.forename: Rohan T
authors.initials: RT
authors.lastname: Ranasinghe
authors.forename: Clive A
authors.initials: CA
authors.lastname: Smith
authors.forename: Shehu M
authors.initials: SM
authors.lastname: Ibrahim
authors.forename: Florian
authors.initials: F
authors.lastname: Hollfelder
authors.forename: Wilhelm T S
authors.initials: WT
authors.lastname: Huck
authors.forename: David
authors.initials: D
authors.lastname: Klenerman
authors.forename: Chris
authors.initials: C
authors.lastname: Abell
mesh_terms: D015415:Biomarkers / Q000032:analysis; D015374:Biosensing Techniques / Q000295:instrumentation*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation*; D008297:Male; D008832:Microchemistry / Q000295:instrumentation*; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D050997:Nanomedicine / Q000295:instrumentation; D017430:Prostate-Specific Antigen / Q000097:blood*; D011471:Prostatic Neoplasms / Q000097:blood* / Q000175:diagnosis*
chemical_list: D015415:Biomarkers; D017430:Prostate-Specific Antigen